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Literature summary for 2.5.1.18 extracted from

  • Federici, L.; Masulli, M.; Di Ilio, C.; Allocati, N.
    Characterization of the hydrophobic substrate-binding site of the bacterial beta class glutathione transferase from Proteus mirabilis (2010), Protein Eng. Des. Sel., 23, 743-750.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression of wild-type and mutant enzymes in Escherichia coli strain XL1-Blue Proteus mirabilis

Protein Variants

Protein Variants Comment Organism
F113A site-directed mutagenesis of the H-site residue, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme Proteus mirabilis
G8A site-directed mutagenesis of the H-site residue Proteus mirabilis
H167A site-directed mutagenesis of the H-site residue, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme Proteus mirabilis
additional information mutations at the H-site can be used to modulate activity at the level of the different catalytic mechanisms operating on the chosen substrates, each mutation showing a different fingerprint, overview Proteus mirabilis
S110A site-directed mutagenesis of the H-site residue, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme Proteus mirabilis
W164A site-directed mutagenesis of the H-site residue the mutant shows altered substrate specificity and activity compared to the wild-type enzyme Proteus mirabilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.453
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant W164A Proteus mirabilis
0.557
-
glutathione pH 6.5, 25°C, mutant H167A Proteus mirabilis
0.577
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant F113A Proteus mirabilis
0.599
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant H167A Proteus mirabilis
0.651
-
glutathione pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
0.653
-
glutathione pH 6.5, 25°C, mutant S110A Proteus mirabilis
0.69
-
glutathione pH 6.5, 25°C, mutant H167A Proteus mirabilis
0.696
-
glutathione pH 6.5, 25°C, mutant S110A Proteus mirabilis
0.732
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant W164A Proteus mirabilis
0.76
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
1.262
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant F113A Proteus mirabilis
1.298
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant S110A Proteus mirabilis
1.393
-
glutathione pH 6.5, 25°C, mutant F113A Proteus mirabilis
1.502
-
glutathione pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
1.863
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
1.95
-
glutathione pH 6.5, 25°C, mutant W164A Proteus mirabilis
2.155
-
glutathione pH 6.5, 25°C, mutant W164A Proteus mirabilis
2.432
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant S110A Proteus mirabilis
3.607
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant H167A Proteus mirabilis
5.509
-
glutathione pH 6.5, 25°C, mutant F113A Proteus mirabilis

Organism

Organism UniProt Comment Textmining
Proteus mirabilis
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from Escherichia coli strain XL1-Blue by anion exchange chromatography, dialysis, ultrafiltration, and chromatofocussing Proteus mirabilis

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.001
-
purified recombinant mutant W164A, pH 6.5, 25°C, substrate 4-nitrobenzyl chloride Proteus mirabilis
0.001
-
purified recombinant wild-type enzyme, pH 6.5, 25°C, substrate 4-nitrobenzyl chloride Proteus mirabilis
0.002
-
purified recombinant mutant H167A, pH 6.5, 25°C, substrate 4-nitrobenzyl chloride Proteus mirabilis
0.004
-
purified recombinant mutant S110a, pH 6.5, 25°C, substrate 4-nitrobenzyl chloride Proteus mirabilis
0.005
-
purified recombinant mutant F113A, pH 6.5, 25°C, substrate 4-nitrobenzyl chloride Proteus mirabilis
0.011
-
purified recombinant mutant F113A, pH 6.5, 25°C, substrate ethacrynic acid Proteus mirabilis
0.017
-
purified recombinant mutant H167A, pH 6.5, 25°C, substrate ethacrynic acid Proteus mirabilis
0.035
-
purified recombinant mutant S110a, pH 6.5, 25°C, substrate ethacrynic acid Proteus mirabilis
0.081
-
purified recombinant wild-type enzyme, pH 6.5, 25°C, substrate ethacrynic acid Proteus mirabilis
0.088
-
purified recombinant mutant W164A, pH 6.5, 25°C, substrate ethacrynic acid Proteus mirabilis
0.17
-
purified recombinant mutant F113A, pH 6.5, 25°C, substrate 1-chloro-2,4-dinitrobenzene Proteus mirabilis
0.48
-
purified recombinant mutant H167A, pH 6.5, 25°C, substrate 1-chloro-2,4-dinitrobenzene Proteus mirabilis
0.67
-
purified recombinant mutant F113A, pH 6.5, 25°C, substrate 1-fluoro-2,4-dinitrobenzene Proteus mirabilis
1.08
-
purified recombinant wild-type enzyme, pH 6.5, 25°C, substrate 1-chloro-2,4-dinitrobenzene Proteus mirabilis
1.31
-
purified recombinant mutant S110a, pH 6.5, 25°C, substrate 1-chloro-2,4-dinitrobenzene Proteus mirabilis
1.91
-
purified recombinant mutant H167A, pH 6.5, 25°C, substrate 1-fluoro-2,4-dinitrobenzene Proteus mirabilis
3.31
-
purified recombinant mutant S110a, pH 6.5, 25°C, substrate 1-fluoro-2,4-dinitrobenzene Proteus mirabilis
3.69
-
purified recombinant wild-type enzyme, pH 6.5, 25°C, substrate 1-fluoro-2,4-dinitrobenzene Proteus mirabilis
7.89
-
purified recombinant mutant W164A, pH 6.5, 25°C, substrate 1-fluoro-2,4-dinitrobenzene Proteus mirabilis
22.69
-
purified recombinant mutant W164A, pH 6.5, 25°C, substrate 1-chloro-2,4-dinitrobenzene Proteus mirabilis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1-chloro-2,4-dinitrobenzene + glutathione 1-chloro-2,4-dinitrobenzene is conjugated through a classic SNAr reaction that proceeds in two steps with the formation of an intermediate Meisenheimer complex Proteus mirabilis S-(2,4-dinitrophenyl)glutathione + HCl
-
?
1-fluoro-2,4-dinitrobenzene + glutathione 1-fluoro-2,4-dinitrobenzene is conjugated through a classic SNAr reaction that proceeds in two steps with the formation of an intermediate Meisenheimer complex Proteus mirabilis S-(2,4-dinitrophenyl)glutathione + HF
-
?
4-nitrobenzyl chloride + glutathione 4-nitrobenzyl chloride is conjugated through a single-step bimolecular nucleophilic substitution (SN2) with a pentacoordinate intermediate Proteus mirabilis S-(4-nitrobenzyl)glutathione + HCl
-
?
ethacrynic acid + glutathione ethacrynic acid is conjugated via a Michael addition reaction through the formation of an enole intermediate Proteus mirabilis ?
-
?

Synonyms

Synonyms Comment Organism
GSH transferase
-
Proteus mirabilis
PmGST
-
Proteus mirabilis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
45
-
mutant F113A Proteus mirabilis
50
-
wild-type GST, and mutants S110A and W164A Proteus mirabilis
60
-
mutant H167A Proteus mirabilis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.12
-
glutathione pH 6.5, 25°C, mutant F113A Proteus mirabilis
0.2
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant F113A Proteus mirabilis
0.43
-
glutathione pH 6.5, 25°C, mutant H167A Proteus mirabilis
0.88
-
glutathione pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
1.03
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant F113A Proteus mirabilis
1.56
-
glutathione pH 6.5, 25°C, mutant S110A Proteus mirabilis
2.13
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
2.59
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant H167A Proteus mirabilis
3.18
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant H167A Proteus mirabilis
4.37
-
glutathione pH 6.5, 25°C, mutant H167A Proteus mirabilis
4.7
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant S110A Proteus mirabilis
6.38
-
glutathione pH 6.5, 25°C, mutant F113A Proteus mirabilis
6.44
-
glutathione pH 6.5, 25°C, mutant S110A Proteus mirabilis
6.78
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
8.66
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant S110A Proteus mirabilis
12.2
-
1-Fluoro-2,4-dinitrobenzene pH 6.5, 25°C, mutant W164A Proteus mirabilis
12.4
-
glutathione pH 6.5, 25°C, wild-type enzyme Proteus mirabilis
14.99
-
glutathione pH 6.5, 25°C, mutant W164A Proteus mirabilis
21.44
-
1-chloro-2,4-dinitrobenzene pH 6.5, 25°C, mutant W164A Proteus mirabilis
29.7
-
glutathione pH 6.5, 25°C, mutant W164A Proteus mirabilis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.5
-
assay at Proteus mirabilis

pH Range

pH Minimum pH Maximum Comment Organism
5 9 activity range, profiles of wild-type and mutant enzymes, overview Proteus mirabilis

General Information

General Information Comment Organism
additional information contribution of different residues in the GSH-binding site to GSH activation, structure analysis of wild-type and mutant enzymes by circular dichroism and fluorospectroscopic analysis, overview Proteus mirabilis